Progress in FY2008 has been in the following areas:[unreadable] [unreadable] 1. STRUCTURAL MODEL FOR THREE-FOLD SYMMETRIC BETA-AMYLOID FIBRILS: In earlier work (Petkova et al., Science 2005), we discovered that the 40-residue beta-amyloid peptide can form more than one fibril structure, that the predominant fibril structure can be determined by subtle variations in growth conditions, that fibrils with distinct morphologies in electron microscope images have distinct underlying molecular structures, and that morphology and molecular structure are self-propagating when pre-existing fibril fragments are used as seeds for the growth of new fibrils. In earlier work (Petkova et al., Biochemistry 2006), we have also developed a full structural model for beta-amyloid fibrils that form under gentle agitation of a beta-amyloid solution and that have mass-per-length (MPL) values, determined by scanning transmission electron microscopy (collaboration with R.D. Leapman, DBEPS, NIBIB), corresponding to two molecular layers in a cross-beta structural motif. We found that fibrils grown under quiescent solution conditions have MPL values that correspond to THREE molecular layers, but our earlier solid state NMR data indicated that two or more quiescent fibril structures might coexist. Now, we have succeeded in preparing highly structurally homogeneous beta-amyloid fibrils with MPL values corresponding to three molecular layers, and we have succeeded in obtaining sufficient structural constraints from solid state NMR to develop a full molecular model. Whereas our previously published model for agitated fibrils had two-fold symmetry about the long fibril axis, the new model (which applies to a different fibril type) has three-fold symmetry. The beta-amyloid peptide conformations in two-fold and three-fold symmetric structures are similar, with essentially the same beta-strand segments and the same in-register parallel beta-sheet tertiary structure, but the conformations of non-beta-strand segments are different, and the contacts between beta-sheets are also different. Structural constraints include extensive NMR, TEM, STEM, AFM, and mutagenesis data. A manuscript describing the new beta-amyloid fibril structure is currently under review.[unreadable] [unreadable] 2. In collaboration with R.B. Wickner's group in NIDDK, we have carried out related studies of amyloid fibrils formed by the yeast prion proteins Rnq1, Sup35p, and Ure2p. In particular, we have shown that Rnq1 fibrils and "scrambled" versions of Sup35p and Ure2p have the same in-register parallel beta-sheet structure as beta-amyloid fibrils.[unreadable] [unreadable] 3. We have initiated studies of beta-amyloid fibrils prepared from synthetic peptide by seeding with amyloid fibrils extracted from brain tissue of deceased Alzheimer's disease patients (collaboration with S.C. Meredith, University of Chicago). Preliminary data indicate that brain-seeded fibrils have a single predominant morphology and molecular structure, and that the structure differs from structures we have examined in previous work on purely synthetic samples. This seeding approach gives us a method for characterizing the molecular structures of in vivo fibrils by solid state NMR and electron microscopy. Future work will seek to identify structural variations among in vivo fibrils that may correlate with neurodegeneration or with localization within the brain.